运用生色基因标记黄瓜根围促生菌(PGPR)筛选菌株

采用三亲交配方法 ,通过Tn7转座系统将lacZY标记基因导入黄瓜根围促生菌 (PG PR)筛选菌株PseudomonasfluorescensCN1 1 6和PseudomonascorrugataCN31的利福平抗性突变株中 ;标记假单胞菌菌株则被赋予了利用乳糖作为唯一碳源的能力 ,在只有乳糖的M9培养基上生长能分解X Gal,菌落显出特有的蓝色 ;经Southern杂交分析 ,证明标记基因lacZY存在于转化菌株的染色体上 ;经验证标记菌株标记性状稳定 ,与对应的野生菌株比较其它性状如培养性状、形态特征、生防效果等基本不变 ;PGPR菌株利福平抗性和生色基因标记的结合 (双标记 )能最大限度地将土壤中引入的PGPR菌株与土著细菌分开 ,检测下限可达 1 0CFU mL ,为PGPR在根围的分子生态学研究提供了一个较好的工具。

INTRODUCTION OF THE CHROMOGENIC GENE TO THE PLANT GROWTH-PROMOTING RHIZOBACTERIA OF CUCUMBER

Using a bicomponent transposition system with the E. coli lacZY gene cloned between Tn7 termini, a sensitive, selectable marker based on expression of the E. coli lac operon genes encoding beta-galactosidase and lactose permease was transformed into the rifampicin resistant mutant of plant growth-promoting rhizobacteria of cucumber, Pseudomonas aeruginosa CN116 and Pseudomonas corrugata CN31, respectively. Transformants were conferred the ability to utilize lactose as a sole carbon source and the ability to cleave the chromogenis substrate X-Gal to show a specific blue color. Southern blotting analysis showed that lacZY gene was inserted into the genome DNA of target strains. Compared with the wild type strains, the cultural characters, morphological features, growth promoting and disease control effects of transformants were almost unchanged, except the new marked phenotype. This marker system enabled the detection of lac+ transformants at sensitivity of 10 CFU/g soil, which makes the further studies on PGPR more easily.